High Penetrance of Pheochromocytoma Associated with the Novel C634Y/Y791F Double Germline Mutation in the RET Protooncogene

Context: Previous studies have shown that double RET mutations may be associated with unusual multiple endocrine neoplasia type 2 (MEN 2) phenotypes. Objective: Our objective was to report the clinical features of patients harboring a previously unreported double mutation of the RET gene and to characterize this mutation in vitro. Patients: Sixteen patients from four unrelated families and harboring the C634Y/Y791F double RET germline mutation were included in the study. Results: Large pheochromocytomas measuring 6.0-14 cm and weighing upto 640 g were identified in the four index cases. Three of the four tumors were bilateral. High penetrance of pheochromocytoma was also seen in the C634Y/Y791F-mutation-positive relatives (seven of nine, 77.7%). Of these, two cases had bilateral tumors, one presented with multifocal tumors, two cases had large tumors (>5 cm), and one case, which was diagnosed with a large (5.5 x 4.5 x 4.0 cm) pheochromocytoma, reported early onset of symptoms of the disease (14 yr old). The overall penetrance of pheochromocytoma was 84.6% (11 of 13). Development of medullary thyroid carcinoma in our patients seemed similar to that observed in patients with codon 634 mutations. Haplotype analysis demonstrated that the mutation did not arise from a common ancestor. In vitro studies showed the double C634Y/Y791F RET receptor was significantly more phosphorylated than either activated wild-type receptor or single C634Y and Y791F RET mutants. Conclusions: Our data suggest that the natural history of the novel C634Y/Y791F double mutation carries a codon 634-like pattern of medullary thyroid carcinoma development, is associated with increased susceptibility to unusually large bilateral pheochromocytomas, and is likely more biologically active than each individual mutation. (J Clin Endocrinol Metab 95: 1318-1327, 2010)

The Absence of Coronary Calcification Does Not Exclude Obstructive Coronary Artery Disease or the Need for Revascularization in Patients Referred for Conventional Coronary Angiography

Objectives This study was designed to evaluate whether the absence of coronary calcium could rule out >= 50% coronary stenosis or the need for revascularization. Background The latest American Heart Association guidelines suggest that a calcium score (CS) of zero might exclude the need for coronary angiography among symptomatic patients. Methods A substudy was made of the CORE64 (Coronary Evaluation Using Multi-Detector Spiral Computed Tomography Angiography Using 64 Detectors) multicenter trial comparing the diagnostic performance of 64-detector computed tomography to conventional angiography. Patients clinically referred for conventional angiography were asked to undergo a CS scan up to 30 days before. Results In all, 291 patients were included, of whom 214 (73%) were male, and the mean age was 59.3 +/- 10.0 years. A total of 14 (5%) patients had low, 218 (75%) had intermediate, and 59 (20%) had high pre-test probability of obstructive coronary artery disease. The overall prevalence of >= 50% stenosis was 56%. A total of 72 patients had CS = 0, among whom 14 (19%) had at least 1 >= 50% stenosis. The overall sensitivity for CS = 0 to predict the absence of >= 50% stenosis was 45%, specificity was 91%, negative predictive value was 68%, and positive predictive value was 81%. Additionally, revascularization was performed in 9 (12.5%) CS = 0 patients within 30 days of the CS. From a total of 383 vessels without any coronary calcification, 47 (12%) presented with >= 50% stenosis; and from a total of 64 totally occluded vessels, 13 (20%) had no calcium. Conclusions The absence of coronary calcification does not exclude obstructive stenosis or the need for revascularization among patients with high enough suspicion of coronary artery disease to be referred for coronary angiography, in contrast with the published recommendations. Total coronary occlusion frequently occurs in the absence of any detectable calcification. (Coronary Evaluation Using Multi-Detector Spiral Computed Tomography Angiography Using 64 Detectors [CORE-64]; NCT00738218) (J Am Coll Cardiol 2010;55:627-34) (C) 2010 by the American College of Cardiology Foundation

Binding and internalization of the melanocyte stimulating hormone receptor ligand [Nle(4), D-Phe(7)]alpha-MSH in B16 melanoma cells

The current study aims to ascertain the fate of the melanocyte stimulating hormone (MSH) receptor and its ligand [Nle(4), D-Phe(7)]alpha-MsH (NDP-MSH) following binding to murine B16 melanoma cells. Cells were incubated with [I-125]-NDP-MSH for up to 180 min and binding, internalization and degradation determined. Intracellular trafficking of the radiolabel was assessed !using Percoll density gradient centrifugation of homogenized cells. Receptor down-regulation and receptor mRNA levels were also measured over 96 hr after exposure to 1 mu M ligand. NDP-MSH accumulation increased with time in a temperature-dependent manner and was inhibited by excess peptide. The ligand was rapidly internalized and translocated to the lysosomal compartment where it was degraded. Internalization was accompanied by a loss or down-regulation of cell surface receptors, suggesting internalization of the NDP-MSH-receptor complex. No recycling of the receptors between the plasma membrane and intracellular compartments could be detected in this cell-hue. Approximately 15% of the surface receptors were resistant to down-regulation, possibly indicating receptor heterogeneity. Down-regulation persisted ibr up to 96 hr and was accompanied by a decrease in MSH receptor mRNA levels 48 hr after treatment. However, before this time, transcript levels were the same in treated and control cells. In contrast to what was seen with NDP-MSH, cell surface receptors removed with trypsin wc:re rapidly replaced. These results show that NDP-MSH not only induced MSH receptor :internalization but also inhibited receptor turnover, resulting in a prolonged down-regulation. It is concluded that, in B16 cells, the MSH receptor undergoes ligand-dependent internalization, resulting in a prolonged down-regulation. Copyright (C) 1996 Elsevier Science Ltd

Can occupational therapy intervention play a part in maintaining independence and quality of life in older people? A randomised controlled trial Jeannine Liddle 1 , 2 , Lyn March 3 , Barbara Carfrae 4 , Terence Finnegan 5 , Jane Druce 6 , Jennifer Schwarz 7 Peter Brooks

The main objective of this study was to see if older people could maintain their quality of life and independence after their homes had been modified and they were using community services as recommended by an occupational therapist. There were 167 study participants aged 69 to 94 years from the Northern Sydney Area, After being assessed at home by an occupational therapist, 105 were randomly allocated to one of two groups, to either have or not have the occupational therapist's recommendations carried out, They were assessed again after six months, A third group did not require any intervention, This group was followed up by telephone and postal questionnaire at six months. The main outcome measures used were the Sickness Impact Profile, the Philadelphia Geriatric Center Morale Scale, the Life Satisfaction Index, assessment of Activities of Daily Living, the Health Assessment Questionnaire and change in residence. After six months there were no difference in outcomes among the three groups. Most study participants remained at a satisfactory level on each measure. Three people had died, One had moved to hostel care and one had moved to a nursing home. A further 14 from the group having no intervention had withdrawn from the study, A secondary objective of this study was to indicate the responsiveness of these outcome measures to change in the short term (over six months) in an elderly population. Twelve-month assessments are in progress and may indicate what to expect from these outcome measures in the medium term.

Development of a synthetic 7-ketocholesterol- rich emulsion resembling chylomicrons

Lipid emulsions that mimic natural lipoproteins help to understand the metabolism and the constitutional organization of circulating lipids. Chylomicrons synthesised by enterocyte cells usually contain oxysterols such as 7-ketocholesterol (7-KC). Here we describe the development of a 7-KC-containing emulsion as a model for oxisterol-rich chylomicron. Different amounts of 7-KC were used. Emulsion characteristics as effective diameter, lipid saturation with radiolabeled lipids was evaluated. In conclusion, the production of a synthetic 7-KC-rich emulsion resembling hylomicrons was feasible, being a model for in vivo metabolism studies.

High sucrose intake in rats is associated with increased ACE2 and angiotensin-(1-7) levels in the adipose tissue

Sucrose-fed rats, a model of metabolic syndrome, are characterized by insulin resistance, obesity, hypertension, and high plasma levels of triacylglycerols and angiotensin II (Ang II). However, whether tissue renin-angiotensin system (RAS) is altered in metabolic syndrome is unclear. To study this issue, food ad libitum and water (C) or 20% sucrose solution (SC) were given to adult male Wistar rats, for 30 days. Body weight (BW), blood pressure (BP), epididymal adipose tissue (EPI) mass, rate of in vivo fatty acid (FA) synthesis in EPI, circulating glucose, insulin, leptin, angiotensins I and II, triacylglycerols, and plasma renin (PRA) and angiotensin-converting enzyme (ACE) activities were evaluated. In kidneys and EPI, gene and protein expression of type 1 (AT(1)) and 2 (AT(2)) Ang II receptors, ACE, angiotensinogen (ACT) as well as protein expression of angiotensin-converting enzyme 2 (ACE2) were determined. In both tissues, Ang I, Ang II and Ang-(1-7) contents were also measured by HPLC. In SC rats higher BP, EPI mass, circulating triacylglycerols, insulin, leptin, PRA and, Ang II were found. In EPI, the rate of in vivo FA synthesis was associated with increased Ang-(1-7), protein expression of AT(1) and AT(2) receptors, ACE2, ACT, and gene expression of ACT although a reduction in ACE activity and in adipose Ang I and Ang II contents was observed. In kidneys, AT(1) and AT(2), ACE and ACT gene and protein expression as well as protein expression of ACE2 were unaltered while Ang II, Ang-(1-7) and ACE activity increased. These RAS component changes seem to be tissue specific and possibly are related to enhancement of FA synthesis, EPI mass and hypertension. (C) 2010 Elsevier B.V. All rights reserved.

Synthetic nanoemulsion resembling a protein-free model of 7-ketocholesterol containing low density lipoprotein: In vitro and in vivo studies

7-ketocholesterol (7-KC) differs from cholesterol by a functional ketone group at C7. It is an oxygenated cholesterol derivative (oxysterol), commonly present in oxidized low-density lipoprotein (LDL). Oxysterols are generated and participate in several physiologic and pathophysiologic processes. For instance, the cytotoxic effects of oxidized LDL have been widely attributed to bioactive compounds like oxysterols. The toxicity is in part due to 7-KC. Here we aimed to demonstrate the possibility of incorporating 7-KC into the synthetic nanoemulsion LDE, which resembles LDL in composition and behavior. This would provide a suitable artificial particle resembling LDL to study 7-KC metabolism. We were able to incorporate 7-KC in several amounts into LDE. The incorporation was evaluated and confirmed by several methods, including gel filtration chromatography, using radiolabeled lipids. The incorporation did not change the main lipid composition characteristics of the new nanoparticle. Particle sizes were also evaluated and did not differ from LDE. In vivo studies were performed by injecting the nanoemulsion into mice. The plasma kinetics and the targeted organs were the same as described for LDE. Therefore, 7-KC-LDE maintains composition, size and some functional characteristics of LDE and could be used in experiments dealing with 7-ketocholesterol metabolism in lipoproteins.

Aerobic Exercise Training-Induced Left Ventricular Hypertrophy Involves Regulatory MicroRNAs, Decreased Angiotensin-Converting Enzyme-Angiotensin II, and Synergistic Regulation of Angiotensin-Converting Enzyme 2-Angiotensin (1-7)

Aerobic exercise training leads to a physiological, nonpathological left ventricular hypertrophy; however, the underlying biochemical and molecular mechanisms of physiological left ventricular hypertrophy are unknown. The role of microRNAs regulating the classic and the novel cardiac renin-angiotensin (Ang) system was studied in trained rats assigned to 3 groups: (1) sedentary; (2) swimming trained with protocol 1 (T1, moderate-volume training); and (3) protocol 2 (T2, high-volume training). Cardiac Ang I levels, Ang-converting enzyme (ACE) activity, and protein expression, as well as Ang II levels, were lower in T1 and T2; however, Ang II type 1 receptor mRNA levels (69% in T1 and 99% in T2) and protein expression (240% in T1 and 300% in T2) increased after training. Ang II type 2 receptor mRNA levels (220%) and protein expression (332%) were shown to be increased in T2. In addition, T1 and T2 were shown to increase ACE2 activity and protein expression and Ang (1-7) levels in the heart. Exercise increased microRNA-27a and 27b, targeting ACE and decreasing microRNA-143 targeting ACE2 in the heart. Left ventricular hypertrophy induced by aerobic training involves microRNA regulation and an increase in cardiac Ang II type 1 receptor without the participation of Ang II. Parallel to this, an increase in ACE2, Ang (1-7), and Ang II type 2 receptor in the heart by exercise suggests that this nonclassic cardiac renin-angiotensin system counteracts the classic cardiac renin-angiotensin system. These findings are consistent with a model in which exercise may induce left ventricular hypertrophy, at least in part, altering the expression of specific microRNAs targeting renin-angiotensin system genes. Together these effects might provide the additional aerobic capacity required by the exercised heart. (Hypertension. 2011;58:182-189.).

Insulin-like growth factor-1 and 17 beta-estradiol down-regulate prostate apoptosis response-4 expression in MCF-7 breast cancer cells

The PKC apoptosis WTI regulator gene, also named prostate apoptosis response-4 (PAR-4), encodes a pro-apoptotic protein that sensitizes cells to numerous apoptotic stimuli. Insulin-like growth factor-1 (IGF-1) and 17 beta-estradiol (E2), two important factors for breast cancer development and progression, have been shown to down-regulate PAR-4 expression and inhibit apoptosis induced by PAR-4 in neuronal cells. In this study, we sought to investigate the mechanisms of regulation of PAR-4 gene expression in MCF-7 cells treated with E2 or IGF-1. E2 (10 nM) and IGF-1 (12.5 nM) each down-regulated PAR-4 expression in MCF-7 cells after 24 h of treatment. The effect of E2 was dependent on ER activation, as demonstrated by an increase in PAR-4 expression when cells were pretreated for 1 h with 1 mu M ICI-182,780 (ICI) before receiving E2 plus ICI. The effect of IGF-1 was abolished by pre-treatment for 1 h with 30 mu M LY294002 (a specific PI3-K inhibitor), and significantly inhibited by 30 mu M SB202190 (a specific p38MAPK inhibitor). We also demonstrated that E2 acts synergistically with IGF-1, resulting in greater down-regulation of PAR-4 mRNA expression compared with E2 or IGF-1 alone. Our results show for the first time that E2 and IGF-1 inhibit PAR-4 gene expression in MCF-7 cells, suggesting that this down-regulation may provide a selective advantage for breast cancer cell survival.

Could the treatment of differentiated thyroid carcinoma with 3.7 and 5.55 GBq of ((131)I)NaI, on an outpatient basis, be safe?

Objectives The first objective of this study was to evaluate the radiological impact on relatives and the environment because of outpatient treatment of differentiated thyroid carcinoma with 3.7 and 5.55 GBq of ((131)I)NaI. The second objective was to determine, analyze, and evaluate whole-body radiation dose to caregivers, the production of contaminated solid waste, and the potentiality of radiation dose and surface contamination existing inside patients` households. Methods Twenty patients were treated on an outpatient basis, taking into consideration their acceptable living conditions, interests, and willingness to comply with medical and radiation-safety guidelines. The caregivers themselves, as well as the potentiality of the radiation dose inside patients` residences, were monitored with a thermo-luminescence dosimeter. Surface contamination and contaminated solid wastes were identified and measured by using a Geiger-Muller detector. Results and discussion Twenty-six monitored individuals received accumulated effective radiation doses of less than 1.0 mSv, and only one 2.8 mSv, throughout the 7 days of measurement. The maximum registered value for the potential of radiation dose inside all living areas was 1.30 mSv. The monitored surface contamination inside patients` dwellings showed a mean value of 4.2 Bq/cm(2) for all surfaces found to be contaminated. A total of 2.5l of contaminated solid waste was generated by the patients with 3.33 MBq of all estimated activity. Conclusion This study revealed that the treatment of differentiated thyroid carcinoma with 3.7 and 5.55 GBq of ((131)I)NaI, on an outpatient basis, can be safe when overseen by qualified professionals and with an adapted radiation-protection guideline. Even considering the radioiodine activity level and the dosimetric methodology applied here, negligible human exposure and a nonmeasurable radiological impact to the human environment were found. Nucl Med Commun 30:533-541 (C) 2009 Wolters Kluwer Health vertical bar Lippincott Williams & Wilkins.

Early short-term versus prolonged low-dose methylprednisolone therapy in acute lung injury

The present study compared the effects of early short-term with prolonged low-dose corticosteroid therapy in acute lung injury (ALI). In total, 120 BALB/c mice were randomly divided into five groups. In the control group, saline was intratracheally (i.t.) instilled. In the ALI group, mice received Escherichia coli lipopolysaccharide (10 mu g i.t.). ALI animals were further randomised into four subgroups to receive saline (0.1 mL i.v.) or methylprednisolone (2 mg center dot kg(-1) i.v.) at 6 h, 24 h or daily (for 7 days, beginning at day 1). At 1, 3 and 8 weeks, in vivo and in vitro lung mechanics and histology (light and electron microscopy), collagen and elastic fibre content, cytokines in bronchoalveolar lavage fluid and the expression of matrix metalloproteinase (MMP)-9 and -2 were measured. In vivo (static elastance and viscoelastic pressure) and in vitro (tissue elastance and resistance) lung mechanics, alveolar collapse, cell infiltration, collagen and elastic fibre content and the expression of MMP-9 and MMP-2 were increased in ALI at 1 week. Methylprednisolone led to a complete resolution of lung mechanics, avoided fibroelastogenesis and the increase in the expression of MMP-9 and MMP-2 independent of steroid treatment design. Thus, early short-term, low-dose methylprednisolone is as effective as prolonged therapy in acute lung injury.

Analysis of human kallikrein 7 expression as a potential biomarker in cervical neoplasia

Overexpression of kallikrein 7, a proteolytic enzyme important for epithelial cell shedding, may be causally involved in carcinogenesis, particularly in tumor metastasis and invasion. In this study, we have evaluated hK7 (human kallikrein 7) protein levels by immunohistochemistry in 367 cervical histological samples including 35 cases of cervicitis, 31 low-grade cervical intraepithelial neoplasia, 51 high-grade cervical intraepithelial neoplasia (H-SIL), 197 squamous cervical carcinomas (SCC) and 53 cervical adenocarcinomas. We have observed that hK7 staining increased with the severity of cervical disease. Intense hK7 staining was found in 15.2% of cervicitis samples, in contrast to 55% of H-SIL and 68% of SCC. Moreover, 92.5% of adenocarcinomas also exhibited intense hK7 staining. Differences in the expression of hK7 could potentially be used as a biomarker for the characterization of different stages of cervical disease.